Ascorbic acid (hereinafter referred to as AsA) is widely used, for example, in the fields of medicines, foodstuffs and cosmetics, but has the disadvantage that it is susceptible to thermal and oxidative degradation. In order to overcome this disadvantage, various derivatives of AsA have been reported. One of the most promising derivatives among them is ascorbic acid-2-phosphate (hereinafter referred to as AsA2P), which shows remarkably high stability in vitro, and is easily converted into AsA in vivo.
Several approaches to synthesize AsA2P by chemical methods have been known (Japanese Published Examined Patent Application Nos. 30328/70, 18191/77 and 15605/73). Chemical phosphorylation of AsA, however, has the problem that various other phosphoric esters of AsA are formed besides the desired AsA2P and that complicated steps for purification of AsA2P are required to eliminate the undesired products. Thus, the chemical phosphorylation of AsA is laborious and expensive.
Compared to the chemical methods microbiological or, enzymatic phosphorylation of AsA (Japanese Published Unexamined Patent Application Nos. 214190/78, 273489/78, 199590/89 and 42996/90) is considered to be more advantageous, in that phosphorylation takes place at a specific site, in this case, at the C-2 position. However, the microorganisms so far proposed for this purpose are of low activity and conversion efficiency of AsA to AsA2P is generally in sufficient for industrial production of AsA2P. Therefore, it has been desired to develop a more efficient process for the preparation of AsA2P.
As a result of extensive studies in process for producing AsA2P, it has now been found that AsA2P can effectively be prepared from AsA and a phosphate donor, by constructing a recombinant DNA comprising a gene isolated from a microorganism belonging to the genus Pseudomonas and coding for AsA phosphorylating enzyme, transforming a microorganism belonging to the genus Escherichia with the recombinant DNA and converting AsA and a phosphate donor into AsA2P by use of the transformed microorganism as an enzyme source.